Cysteinylation of MHC Class II Ligands: Peptide Endocytosis and Reduction Within APC Influences T Cell Recognition

Abstract

Peptides bind cell surface MHC class II proteins to yield complexes capable of activating CD4+ T cells. By contrast, protein Ags require internalization and processing by APC before functional presentation. Here, T cell recognition of a short peptide in the context of class II proteins occurred only after delivery of this ligand to mature endosomal/lysosomal compartments within APC. Functional and biochemical studies revealed that a central cysteine within the peptide was cysteinylated, perturbing T cell recognition of this epitope. Internalization and processing of the modified epitope by APC, was required to restore T cell recognition. Peptide cysteinylation and reduction could occur rapidly and reversibly before MHC binding. Cysteinylation did not disrupt peptide binding to class II molecules, rather the modified peptide displayed an enhanced affinity for MHC at neutral pH. However, once the peptide was bound to class II proteins, oxidation or reduction of cysteine residues was severely limited. Cysteinylation has been shown to radically influence T cell responses to MHC class I ligands. The ability of professional APC to reductively cleave this peptide modification presumably evolved to circumvent a similar problem in MHC class II ligand recognition.

HLA‐DM negatively regulates HLA‐DR4‐restricted collagen pathogenic peptide presentation and T cell recognition

“…Human B lymphoblastoid cell lines 6.16 and BLS-1, and the human monocytes THP-1 were cultured in complete IMDM with 10% heat-inactivated fetal bovine serum, 50 U/mL penicillin and 50 lg/mL streptomycin [13,36,40,45]. 6.16, BLS-1 and THP-1 Eur.…”

“…Cells were subsequently washed twice in PBS and fixed with 1% paraformaldehyde prior to cultivation with CII or jII peptide-specific T cells 4027/99 and 1.21, respectively. T cell production of IL-2 was measured using HT-2 cells [40,50]. All assays were repeated at least three times and the data were corrected for isotope counting efficiency and expressed as mean corrected cpm (ccpm) + SEM.…”

“…6A and C). When live or prefixed APC were incubated with a control peptide j 145-159 , which does not require internalization and processing for T cell recognition [40], no changes in peptide presentation as monitored by T cell proliferation were observed ( Fig. 6B and D).…”

“…T cell hybridomas and the IL-2-dependent cell line HT-2 were cultured in RPMI 1640 with 10% FBS, 50 U/mL penicillin, 50 lg/mL streptomycin and 50 lM b-mercaptoethanol. For HT-2 cells, 20% Con A supernatant (T-STIM; Collaborative Biomedical Research, Bedford, MA) was added as a supplement [37,40]. …”

“…For HT-2 cells, 20% Con A supernatant (T-STIM; Collaborative Biomedical Research, Bedford, MA) was added as a supplement [37,40].…”